This application is based on and claims priority of Japanese Patent Application No. Hei 11-270297 filed on Sep. 24, 1999, the content of which is incorporated herein by reference.
1. Field of the Invention
The present invention relates to a focusing system for a microscope in general and especially relates to a focusing system for an incident-light fluorescence microscope.
2. Description of the Related Art
Traditionally, in the field of microscopy, in which a microscope observes an optical image of a sample transmitted through an objective lens on a video monitor, and takes a picture of the optical image, an auto focusing system to detect a focal point of the objective lens, and to adjust the focal point to an image plane of a TV camera and a camera film is one of most important elements present.
Recently, an incident-light fluorescence microscopes to observe fluorescent light radiated from a sample, for example, biopsy tissue and biological cells, dyed by a fluorescent reagent by exciting the fluorescent reagent have become popular. Now, auto focusing systems are expected to operate with the incident-light fluorescence microscope.
An auto focusing system used in substantially this kind of fluorescence observation is disclosed in Japanese Laid-Open Patent Publication No. Hei 9-189849. Japanese Laid-Open Patent Publication No. Hei 9-189849 teaches a method to correct a focal shift on the basis of chromatic aberration coming from each imaging optical system of a photo system and a focusing system by using a wavelength of fluorescence.
However, the conventional auto focusing system for the incident-light fluorescence microscope has a problem in that the amount of light of an observed fluorescent wavelength is so small, that it is necessary to spend more time storing incident light until the amount of incident light is adapted to a range of a photo acceptance unit used to detect a focal point. That is, the problem is that the processing time for an automatic focus is overlong.
The above problem causes low efficiency of observation. Furthermore, excitation light from the upper side of the sample is continuously irradiated on the sample during an automatic focus operation, so that the sample is forced to needlessly be subjected to fluorescence photo-bleaching leading to discoloration. Discoloration causes a fatal problem in fluorescence observation.
The present invention provides a focusing system for a microscope and a reflected illumination fluorescence microscope using the focusing system which overcome these problems. It has an objective lens, a sample stage, a reflected illumination system for generating fluorescence from a sample, a transmitted illumination system for irradiating light on the sample to capture a transmitted optical image, a set of optical elements for forming the transmitted optical image on the basis of phase information included in light transmitted through the sample, an optical element for dividing the fluorescence image and the transmitted optical image, a sensor for capturing the transmitted optical image divided by the optical element for dividing light, a focus detecting section for detecting a focusing level of the transmitted optical image on the basis of a signal output from the sensor, and a driver for moving at least one of the objective lens and the stage to focus on the sample on the basis of the focusing level.